Regulation of the Na/K Pump by RNA Editing

RNA 编辑对 Na/K 泵的调节

基本信息

批准号：
7885009
负责人：
JOSHUA J.C. ROSENTHAL
金额：
$ 29.71万
依托单位：
UNIVERSITY OF PUERTO RICO MED SCIENCES
依托单位国家：
美国
项目类别：
财政年份：
2010
资助国家：
美国
起止时间：
2010-08-01 至 2015-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7885009
关键词：
ATP phosphohydrolase Abbreviations Action Potentials Adenosine Affect Affinity Amino Acids Antisense RNA Arrhythmia Axon Back Binding Binding Sites Bioinformatics Ca(2+)-Transporting ATPase Cardiac Cell membrane Cells Codon Nucleotides DRADA2b protein Data Dependence Depressed mood Disease Double-Stranded RNA Binding Domain Dystonia 12 Endoplasmic Reticulum Enzymes Equilibrium Familial Hemiplegic Migraine Fiber Financial compensation Future GTP-Binding Proteins Ganglia Genes Genetic Glutamate Receptor Goals Hereditary Disease Human Hydrophobicity Ions Lead Learning Letters Link Lobe Membrane Part Membrane Potentials Messenger RNA Methods Modeling Molecular Molecular Conformation Mutation Myxoid cyst N Domain Nature Nervous system structure Neuraxis Nomenclature Nucleotides Oligonucleotides Optic Lobe Pathology Peripheral Permeability Phosphorylation Physiological Phytic Acid Play Population Positioning Attribute Potassium Channel Process Protein Isoforms Proteins Public Health Publishing Pump RNA Editing Regulation Research Role Shotguns Signal Transduction Site Squid Steroids Structure Structure of stellate ganglion Surface Synapses System Tertiary Protein Structure Testing Time Trefoil Motif Vertebrates Work base digoxin receptor dsRNA adenosine deaminase extracellular gain of function mutant neuronal cell body null mutation public health relevance receptor binding relating to nervous system serotonin receptor solute therapeutic development tool voltage

项目摘要

DESCRIPTION (provided by applicant): This project focuses on how the Na/K pump is regulated by RNA editing. A central premise to our approach is that naturally occurring codon changes, caused by RNA editing, can lead us to functionally important regions of the Na/K pump. They might also be used as tools to compensate for rapid-onset dystonia parkinsonism, genetic disease associated with the human Na/Ka3 subunit. The squid nervous system will be used as a model because RNA editing is extensive in cephalopods and because the editing sites that we have identified cause a gain of function, a rare phenomenon for a mutation. Preliminary data show that Na/K pump mRNAs can be edited at three codons, two in the phosphorylation domain (P), and the other at the top of the 7th transmembrane span (M7). These changes affect critical components of the pumping cycle. For example, the edits in the P domain increase the apparent affinity for ATP and the edit in M7 regulates how Na is released to the outside. Using an electrophysiological approach, the proposal's first two aims will characterize the mechanism by which these edits exert their function. The goal of the last aim is to see whether squid RNA edits can compensate for depressed turnover rates in human Na/Ka3 caused by mutations associated with rapid-onset dystonia parkinsonism. In addition, not only will we study if they can compensate, we will also try to develop a method to introduce these edits into human pumps at the level of mRNA. Preliminary data show that the human editing enzyme ADAR2 is capable of editing squid mRNAs because it recognizes the appropriate secondary structure. We hypothesize that we can trick human ADAR2 into editing human pumps by mimicking the squid secondary structure with an antisense RNA oligo. From the standpoint of public health, this work is significant on several fronts. The Na/K pump creates the ion gradient that is required for excitability and the majority of solute transport across cell membranes. With the first crystal structure recently published, this is an opportune time to learn more about how the Na/K pump operates. Clinically, the Na/K pump is important because it is the receptor of digoxin, a widely prescribed cardiac steroid used to control many cardiac arrhythmias. Further, two neural disorders have been directly correlated with mutations to the Na/K pump: familial hemiplegic migraine, which is linked to the Na/Ka2 subunit, and rapid-onset dystonia parkinsonism, which is linked to the Na/Ka3 subunit. Results from this proposal will be directly relevant to the development of therapeutics for rapid-onset dystonia parkinsonism. The general approach may also prove relevant for a wide variety of genetic disorders. PUBLIC HEALTH RELEVANCE: The Na/K pump plays a vital role in establishing ion gradients across cells. Preliminary data shows that its ability to pump can be regulated by RNA editing. This proposal focuses on understanding how RNA editing regulates the Na/K pump and how it might be used to treat genetic disorders that affect the Na/K pump.

描述（由申请人提供）：该项目重点研究 Na/K 泵如何通过 RNA 编辑进行调节。我们方法的一个核心前提是，由 RNA 编辑引起的自然发生的密码子变化可以引导我们到达 Na/K 泵的功能重要区域。它们还可以用作补偿快速发作的肌张力障碍帕金森症（与人类 Na/Ka3 亚基相关的遗传病）的工具。鱿鱼神经系统将被用作模型，因为 RNA 编辑在头足类动物中广泛存在，而且我们已经确定的编辑位点会导致功能获得，这是一种罕见的突变现象。初步数据显示，Na/K 泵 mRNA 可以在三个密码子处进行编辑，其中两个位于磷酸化结构域 (P)，另一个位于第七次跨膜跨度 (M7) 的顶部。这些变化会影响泵循环的关键组成部分。例如，P 结构域中的编辑增加了对 ATP 的表观亲和力，M7 中的编辑调节了 Na 向外部释放的方式。使用电生理学方法，该提案的前两个目标将描述这些编辑发挥其功能的机制。最后一个目标是看看鱿鱼 RNA 编辑是否可以补偿由与快速发作的肌张力障碍帕金森症相关的突变引起的人类 Na/Ka3 周转率下降。此外，我们不仅会研究它们是否能够补偿，我们还将尝试开发一种方法，在 mRNA 水平上将这些编辑引入人类泵中。初步数据表明，人类编辑酶 ADAR2 能够编辑鱿鱼 mRNA，因为它可以识别适当的二级结构。我们假设我们可以通过用反义 RNA 寡核苷酸模拟鱿鱼的二级结构来欺骗人类 ADAR2 来编辑人类泵。从公共卫生的角度来看，这项工作在几个方面都具有重要意义。 Na/K 泵产生兴奋性和大部分溶质跨细胞膜运输所需的离子梯度。随着最近发布的第一个晶体结构，现在是了解更多有关 Na/K 泵如何运行的好时机。在临床上，钠/钾泵很重要，因为它是地高辛的受体，地高辛是一种广泛使用的心脏类固醇，用于控制许多心律失常。此外，两种神经疾病与 Na/K 泵突变直接相关：家族性偏瘫偏头痛（与 Na/Ka2 亚基相关）和快速发作的肌张力障碍帕金森症（与 Na/Ka3 亚基相关）。该提案的结果将与快速发作的肌张力障碍帕金森症的治疗方法的开发直接相关。一般方法也可能与多种遗传性疾病相关。公共健康相关性：Na/K 泵在建立细胞间离子梯度方面发挥着至关重要的作用。初步数据表明，其泵送能力可以通过RNA编辑来调节。该提案的重点是了解 RNA 编辑如何调节 Na/K 泵，以及如何使用它来治疗影响 Na/K 泵的遗传性疾病。